The use of thermostable and preferably thermophilic enzymes is highly desired in industry as many enzymatic industrial reactions are performed at high temperatures. Examples include, but are not limited to, the use of peptidases in sectors including, but not limited to, food production and processing such as e.g. cereal processing and baking (e.g. production of bread, pastry), the feed industry (e.g. pet food production), the detergent industry (e.g. laundry detergents), the textile industry (e.g. biopolishing of wool and silk, silk degumming), etc. However, some enzymes induce allergenicity and/or other health issues at ambient temperatures. The application of these enzymes has to be performed with great care. It would therefore be desirable in these and other applications to inactivate the enzyme at lower temperature without compromising the enzyme activity at elevated temperatures. Also, in certain circumstances, for example in food processing, it is undesirable to have enzymes acting at low temperatures, e.g. at room temperature, when other processing steps occur. During these first processing steps enzyme activity is not desired, while in later processing steps at elevated temperature the enzyme activity may be beneficial for the end product quality.
In general, bringing enzyme-inhibitor complexes at higher temperatures simultaneously denatures both, the inhibitor and the enzyme, which does not result in regaining enzymatic activity. Upon heat treatment of the complex between carrot pectin methyl esterase (PME) and its inhibitor (PMEI) from kiwi, enzyme and inhibitor were not driven apart. Yet, the complex denatured (and aggregated) as one entity, following first-order kinetics. Upon pressure treatment, above 300 MPa, the difference in reaction rate of carrot PME with and without PMEI gradually diminished, pointing to a declining amount of inhibited PME. This tendency indicates a pressure-induced dissociation of the PME-PMEI complex. While the liberated kiwi PMEI may be partly inactivated irreversibly, the liberated carrot PME behaves similarly to the other PME present, i.e., a putative reversible inactivation occurs (Jolie et al. (2009) Innovative Food Science & Emerging Technologies 10(4): 601-609). These findings were supported by size exclusion chromatography studies in which the behaviour of the PME-PMEI complex at elevated temperature or pressure levels was determined. Heat treatment proved not to dissociate the complex, but rather to denature the complex as one entity. In contrast, high pressure treatment induced disunion of enzyme and inhibitor, followed by gradual inactivation of PME and PMEI (Jolie et al. (2009) Journal of Agricultural and Food Chemistry 57(23): 11218-11225).
It is an object of the present invention to provide enzyme preparations which are inactive at lower temperatures, but active at elevated temperatures. The use of such enzyme preparations can advantageously reduce or abolish potential health issues associated with the (industrial) application of the enzymes at ambient temperature. Also, in certain applications, the use of enzyme preparations which are inactive at lower temperatures, but active at elevated temperatures, has a beneficial effect on the overall process quality and/or on the quality of the end product (storage, texture, etc.).